论文部分内容阅读
【目的】获得具有结肠靶向的纳米载体。【方法】采用SOE-PCR方法将具有结肠靶向的TK肽序列插入到猪细小病毒(PPV)结构蛋白VP2的环2和环4区域得到TK-vp2(?vp2)基因,在Bac-to-Bac?杆状病毒表达系统中构建、表达和自组装。【结果】通过SOE-PCR方法扩增获得?vp2基因,在Bac-to-Bac?杆状病毒表达系统中构建得到Bacmid-?vp2,经脂质体转染至Sf9昆虫细胞得到重组杆状病毒。直接免疫荧光试验、SDS-PAGE和Western blot检测结果表明?VP2蛋白在Bac-to-Bac?杆状病毒表达系统中获得融合表达,目的蛋白约70 k D;透射电子显微镜结果显示?VP2能自组装形成病毒样颗粒(TK-VLPs),直径范围在22 nm-30 nm。【结论】获得纳米载体TK-VLPs,为进一步研究其作为结肠靶向的纳米载体奠定物质基础。
【Objective】 To obtain colon-targeted nanocarriers. 【Method】 The TK-vp2 (? Vp2) gene was amplified by SOE-PCR and inserted into the loop 2 and loop 4 of the VP2 protein of porcine parvovirus (PPV) Bac? Baculovirus expression system construction, expression and self-assembly. 【Result】 The gene vp2 was amplified by SOE-PCR and Bacmid-vp2 was constructed by Bac-to-Bac baculovirus expression system. The recombinant baculovirus was transfected into Sf9 insect cells by liposome. . Direct immunofluorescence assay, SDS-PAGE and Western blot results showed that VP2 protein was expressed in Bac-to-Bac baculovirus expression system, the target protein was about 70 kD. Transmission electron microscopy showed that VP2 could The assembled virus-like particles (TK-VLPs) range in diameter from 22 nm to 30 nm. 【Conclusion】 The nanocarriers TK-VLPs were obtained and laid the material foundation for further study on their use as colon-targeted nanocarriers.